A considerable number of gastric cancers derive from stomach mucosa where chronic atrophic gastritis is severe and extensive. Based on the fact that the serum pepsinogen levels provide a precise measure of the extent of chronic atrophic gastritis, we have devised a mass screening method involving serum pepsinogen measurement to identify subjects at high risk of gastric cancer. In 1991, we screened 4,647 workers (male: 4,113, female: 534, mean age: 49.0 years) at a Japanese company using this method. Out of 875 subjects (18.8%) with a serum pepsinogen I level of less than 50 μg/liter and a pepsinogen I/II ratio of less than 3.0, 676 subjects (14.5%) were selected for further investigation by endoscopy. This led to the detection of four subjects (0.086%) with gastric cancer (three in an early stage) and four subjects with adenoma. The cancer detection rate of this new screening method was comparable, and in some respects superior, to that of the traditional barium X‐ray screening. Since the incidence of test‐positive subjects was as low as 10% amongst subjects aged less than 40, this screening method appears to be especially useful for screening of younger generations. The new method is less expensive than the traditional barium X‐ray and subjects experience little discomfort. Further, many serum samples can be quickly measured simultaneously. The results of this study have indicated that serum pepsinogen screening provides a valuable method for detecting gastric cancers.

To understand the role of endogenous AP-1 activity in cellular transformation induced by oncogenes, we have made use of a fos mutant (supfos-1) and a jun mutant (supjun-1), either of which can function as a transdominant inhibitor of AP-1-mediated transcriptional regulation. Chicken embryo fibroblasts (CEF) infected with a series of transforming retroviruses were doubly infected with retrovirus carrying supfos-1 or supjun-1, and suppression of cellular transformation was monitored in terms of reversion to normal cellular morphology or acquisition of anchorage-dependent growth. Cellular transformation induced by several exogenously expressed transforming genes of thefos orjun family was efficiently suppressed, as expected. CEF transformed by v-src, v-yes, v-.fps, c-Ha-ras, and N-terminally truncated c-rafwere also induced to revert to the normal phenotype by these transdominant mutants, suggesting that functional transcription factor AP-1 activity is essential for the cellular transformation induced by these oncogenes. The suppression is not attributable to nonspecific inhibition of cellular proliferation, because CEF transformed by v-ros or v-myc were not induced to revert to the normal phenotype. We next analyzed changes in all known components of chicken AP-1 induced by v-src, c-Ha-ras, or activated c-raf transformation. The levels of both Fra-2 and c-Jun expression were elevated two-to fourfold, and hyperphosphorylation of Fra-2 was also observed. We further showed that Fra-2-c-Jun heterodimer is mainly responsible for the elevated AP-1 DNA-binding activity in these transformed cells, and we propose that this heterodimer play a crucial role in the transformation induced by these oncogenes.

We have analyzed the transcriptional regulation of the fra-2 gene in chicken embryo fibroblasts. Like c-fos, fra-2 was inducible by phorbol ester, cAMP and calcium ionophore, as well as serum. In all three cases, the induction of two species of fra-2 transcript (5.7 kb and 6.8 kb) was delayed and prolonged compared with that of c-fos mRNA. The size difference between the two transcripts was attributable to the heterogeneity of the 3'-end, probably reflecting utilization of different polyadenylation sites. The major transcriptional start point is located at 30 bp downstream of a TATA-like sequence. In the fra-2 promoter region, which is located in a typical CpG island, enhancer consensus sequences such as SCM, SRE, GC boxes and CRE-like sequences were detected upstream of the TATA-like sequence in the same order as that in the 5'-upstream region of the chicken c-fos gene. Fibroblast transfection studies with a series of promoter deletion constructs positioned upstream of bacterial chloramphenicol acetyltransferase indicated, however, that SRE-like sequence is not the sole responsible element for the serum induction, and that a minimal fragment containing no SRE-like sequence is sufficient for this induction. Two typical AP-1 sequences are located between the major transcriptional initiation site and the coding sequence, and the binding activity of protein complexes to these sequences was induced by serum.

Although a replication-competent retrovirus that carries junD has no transforming activity in chicken embryo fibroblasts, we have isolated mutant viruses that have spontaneously acquired transforming activity. The molecularly cloned junD genes of three such mutant viruses (T1, T2, and T3) were shown to be responsible for the cellular transformation. DNA sequence analysis indicated that a specific polynucleotide in the junD sequence was tandemly multiplied three times of five times in T1 and T2, respectively. The repeated polynucleotide encodes 16 amino acid residues that are located in a highly conserved region among Jun family proteins. The junD mutation in T3 involved an inversion, a translocation, and nucleotide substitutions that caused drastic amino acid exchanges in another well-conserved region among Jun family proteins. The transcriptional activity of these mutants was analyzed by means of transient expression experiments in F9 cells using a reporter gene containing a single AP-1 binding site. Compared with the wild-type JunD, none of them showed enhanced transactivating activity in the forms of homodimers or of heterodimers with c-Fos or Fra-1. However, they did exhibit much higher transactivating activity than the wild type when they formed heterodimers with Fra-2, indicating that the mutated regions function as transactivation domains in a partner-specific manner. Since we have previously reported that there is a basal level of Fra-2 expression in chicken embryo fibroblasts, the results may indicate that protein complexes between JunD mutants and Fra-2 play a crucial role in the cellular transforming activity.

Undifferentiated glandular stomach tissue fragments from 16.5-day fetal rats were transplanted under the kidney capsule of syngeneic adult rats, and the proliferation, differentiation and morphogenesis of the transplanted tissues were investigated. Gastric epithelial cells began to invaginate 3-4 days after the transplantation and immature glands were formed after 1 week. During the period, there was a gradual increase in the expression of pepsinogen and cathepsin E, markers of cytodifferentiation of the stomach epithelia, both at protein and mRNA levels. Cathepsin E was weakly expressed in undifferentiated gastric epithelial cells at 16.5 days of gestation, and a higher level of the expression was observed in differentiated epithelia of the transplants. In contrast, the pepsinogen-producing cells first appeared around days 3-4 after transplantation and gradually increased in number to about 30% of the epithelial cells and became localized at the bottom of the gland. During the period of the experiment up to 1 month, the pepsinogen-producing cells were all positive for class III mucin and cathepsin E, indicating the immature character of these cells. In addition, no parietal cells were observed. When the tissue fragments were transplanted into adrenalectomized animals, the epithelial differentiation and morphogenesis was suppressed, but its proliferation was enhanced. The observed changes were reversed by hydrocortisone replacement. These results suggest that the development of the 16.5-day fetal stomach is regulated intrinsically to a certain extent by the genetic program of the cells involved and various gastric functions develop in the absence of luminal stimulation, stage-specific systemic hormonal change, neuronal regulation or other systemic influences, and that glucocorticoids modulate the developmental program of the fetal stomach tissues.